Ehrlichia-induced hemophagocytic lymphohistiocytosis in a pediatric kidney transplant recipient.

BACKGROUND: Kidney transplant patients are susceptible to a variety of infections in the post-transplant period due to the use of immunosuppressant medications. Ehrlichiosis is a rare infection in solid organ transplant recipients with signs and symptoms that mimic rejection and other viral infections. Hemophagocytic lymphohistiocytosis (HLH) is a potentially fatal hyperinflammatory syndrome that can be triggered by infections. METHODS: We describe a pediatric kidney transplant recipient who experienced secondary HLH due to ehrlichiosis within the initial post-transplant month. RESULT: Our patient improved after treatment with doxycycline, corticosteroids, and intravenous immunoglobulin (IVIG). CONCLUSION: Clinicians should consider infections such as ehrlichiosis as a potential cause of illness in febrile solid organ transplant recipients in immediate post-transplant period, especially when accompanied by a compatible exposure history.

Assessment of Risk Factors and Outcomes of Severe Ehrlichiosis Infection.

Importance: Ehrlichiosis cases in the US have increased more than 8-fold since 2000. Up to 57% of patients with ehrlichiosis require hospitalization and 11% develop a life-threatening complication; however, risk factors for serious disease are not well documented. Objective: To examine risk factors associated with severe ehrlichiosis. Design, Setting, and Participants: An analytic cross-sectional study of patients diagnosed with ehrlichiosis by polymerase chain reaction (PCR) between January 1, 2007, and December 31, 2017, was conducted in a single tertiary-care center in a region endemic for ehrlichiosis. Analysis was performed from February 27, 2018, to September 9, 2020. A total of 407 positive Ehrlichia PCR results were identified from 383 unique patients, with 155 unique patients meeting study criteria. Patients hospitalized at other institutions who had a positive Ehrlichia PCR performed as a reference test (n = 222) were excluded as no clinical data were available. Electronic medical record review was performed to collect demographic, clinical, laboratory, treatment, and outcomes data. Cases were excluded when there were insufficient clinical data to assess the severity of illness (n = 3) and when the clinical illness did not meet the case definition for ehrlichiosis (n = 3). Exposures: Date of presentation, onset of symptoms, date of PCR testing, date of treatment initiation, site of care, age, birth sex, race/ethnicity, Charlson Comorbidity Index, trimethoprim with sulfamethoxazole use within the prior 2 weeks, and immunosuppression. Main Outcomes and Measures: Requirement for intensive care unit (ICU) admission. Results: Of the 155 patients who met inclusion criteria, 99 patients (63.9%) were men, and 145 patients (93.5%) identified as non-Hispanic White; median age was 50 years (interquartile range, 23-64 years). Intensive care unit admission was indicated in 43 patients (27.7%), 94 patients (60.6%) were hospitalized on general medical floors, and 18 patients (11.6%) received care as outpatients. In adjusted analysis, time to treatment initiation was independently associated with an increased risk for ICU admission (adjusted prevalence ratio [aPR], 1.09; 95% CI, 1.04-1.14; P < .001). Documentation of tick exposure was independently associated with a decreased risk for ICU admission (aPR, 0.54; 95% CI, 0.34-0.86; P = .01). There appeared to be a nonsignificant change toward a decreased need for ICU care among immunosuppressed persons (aPR, 0.51; 95% CI, 0.26-1.00; P = .05). Conclusions and Relevance: This study suggests that delay in initiation of doxycycline therapy is a significant factor associated with severe ehrlichiosis. Increased recognition of infection by front-line clinicians to promote early treatment may improve outcomes associated with this increasingly common and life-threatening infection.

Case Report: Anaplasmosis in Canada: Locally Acquired Anaplasma phagocytophilum Infection in Alberta.

Human granulocytic anaplasmosis is an obligate intra-granulocytic parasite that is transmitted by Ixodes scapularis and Ixodes pacificus in North America. We report on the second laboratory-confirmed case of Anaplasma phagocytophilum acquired within the province of Alberta, Canada. A 67-year-old woman from the Edmonton health zone developed nonspecific systemic symptoms including fatigue, night sweats, myalgia, headaches, and fever 6 days after noticing a tick on her left upper arm in May of 2017 (day 0). On day 13, she was found to have thrombocytopenia. Her symptoms progressed until day 16 when she was treated empirically with doxycycline, at which time she slowly improved over the subsequent 2 months. The tick was later identified as a partially engorged female blacklegged tick, I. scapularis, and it was positive for A. phagocytophilum DNA by PCR. Anaplasma serology performed retrospectively on blood samples collected on days 13, 31, and 52 showed a greater than 4-fold increase in A. phagocytophilum (IgG titers from less than 1:64 on day 13 to 1:2048 on days 31 and 52), consistent with an acute infection. Although populations of blacklegged ticks are not yet established in Alberta, suspicion should remain for tick-borne diseases because infected ticks are introduced into the province by migrating birds. This case report highlights the need to remind physicians and other public health professionals that rare, non-endemic tick-borne diseases can occasionally occur in low-risk jurisdictions.

Infection by Anaplasma phagocytophilum Requires Recruitment of Low-Density Lipoprotein Cholesterol by Flotillins.

Anaplasma phagocytophilum is an obligatory intracellular bacterium that proliferates in membrane-bound inclusions. A. phagocytophilum is dependent on cholesterol and acquire cholesterol from low-density lipoprotein (LDL) endocytosed by mammalian host cells. The mechanism of cholesterol transport to Anaplasma inclusions, however, is not fully understood. Flotillin-1 (FLOT1) and FLOT2 are cholesterol-associated membrane proteins that form a heterodimer and/or oligomer complex. Here, we found that Anaplasma infection was significantly reduced by small interfering RNA (siRNA) knockdown of FLOT1 or FLOT2. Anaplasma inclusions were encircled with small vesicles containing endogenous FLOT1 or FLOT2 or with ectopically expressed FLOT1-mCherry and FLOT2-green fluorescent protein (FLOT2-GFP). FLOT1- and FLOT2-containing vesicles were enriched with unesterified cholesterol, as indicated by labeling with filipin and aminomethyl coumarin acetic acid-conjugated theonellamide. Localization of FLOT2 to Anaplasma inclusions was dependent on cholesterol, as FLOT2-GFP bearing two mutations in the cholesterol recognition/interaction motif could not target the inclusions. The cholesterol-sequestering agent methyl-ß-cyclodextrin abrogated FLOT1 localization to Anaplasma inclusions and cleared infection. FLOT2-GFP also localized to fluorescent 3,3'-dioctadecylindocarbocyanine (DiI)-LDL-containing vesicles, including those surrounding Anaplasma inclusions. FLOT2 siRNA knockdown blocked DiI-LDL trafficking to Anaplasma inclusions and reduced bacteria-associated cholesterol amount, and therefore inhibiting Anaplasma infection. Vesicles containing acid lipase, which hydrolyzes LDL cholesterol esters to free cholesterol, colocalized with FLOT2 and encircled Anaplasma inclusions, while the acid lipase inhibitor orlistat significantly inhibited Anaplasma replication. Together, the data revealed that FLOTs are crucial for Anaplasma replication in host cells, likely by aiding vesicular traffic of LDL-derived free cholesterol to Anaplasma inclusions, and suggest a new way of inhibiting Anaplasma infection.IMPORTANCE Cholesterol is essential for animal cells, but most bacteria do not depend on cholesterol and instead lack cholesterol. However, the intracellular Gram-negative bacterium Anaplasma phagocytophilum that causes human granulocytic anaplasmosis (HGA) is unusual, as it contains significant amount of cholesterol and depends on cholesterol for survival and infection. A. phagocytophilum lacks genes for cholesterol biosynthesis or modification but acquire cholesterol from host cells exclusively from the LDL uptake pathway by a yet-to-be defined mechanism. Here, we uncovered a role of cholesterol-binding proteins FLOT1 and FLOT2 in LDL-derived cholesterol trafficking to Anaplasma inclusions and cholesterol acquisition by Anaplasma species. Importantly, we found that FLOTs localize to A. phagocytophilum-containing inclusions and the compartments containing LDL, and the acid lipase inhibitor orlistat significantly inhibits Anaplasma replication. Our data suggest a fundamental role of FLOTs in intracellular vesicular transport of LDL-derived free cholesterol and may provide insight regarding a new therapeutic target for HGA treatment.

Cerebral haemorrhage as a clinical manifestation of human ehrlichiosis.

A 16-year-old young man presented to the emergency room with new-onset generalised tonic-clonic seizures. Examination showed a Glasgow score of 13 and predominantly crural left hemiparesis. Imaging demonstrated a right frontoparietal haemorrhage of non-vascular origin with perilesional oedema. Surgical drainage was carried out, but rebleeding occurred within 24 hours following surgery, and again 1 week after discharge. On reinterrogation and examination, Ehrlichia canis infection was suspected and empirical management with doxycycline was begun. Improvement was evident 72 hours after antibiotic initiation, and PCR confirmed the diagnosis; thus, doxycycline was continued for 6 months. After 2 years, seizures recurred and treatment was reinstated with good clinical response. However, seizures reappeared whenever treatment discontinuation was attempted. Lacking alternatives, doxycycline was maintained up to the third year following the initial episode. Subsequently, the patient showed complete resolution without neurological sequelae up to his last follow-up visit, 12 months following treatment cessation.

Ehrlichiosis Induced Aseptic Meningitis and Second-Degree Heart Block in an Adolescent Male.

We present a previously healthy 16-year-old male with ehrlichiosis-induced aseptic meningitis and subsequent second-degree heart block.

Butyrylcholinesterase as a marker of inflammation and liver injury in the acute and subclinical phases of canine ehrlichiosis.

The aim of this study was to evaluate the role of butyrylcholinesterase (BChE) as a marker of inflammation and liver injury in the acute and subclinical phases of canine ehrlichiosis. Forty-two serum samples of dogs naturally infected with Ehrlichia canis were used, of which 24 were from animals with the acute phase of the disease and 18 with subclinical disease. In addition, sera from 17 healthy dogs were used as negative controls. The hematocrit, BChE activity, hepatic injury (alanine aminotransferase (ALT) and aspartate aminotransferase (AST)), nitric oxide, and cytokines levels were evaluated. The BChE activity was significantly elevated (P<0.05) in dogs with the acute phase of the disease when compared to healthy animals. However, there was a reduction on BChE activity on dogs with subclinical disease compared to the other two groups. AST and ALT levels were significantly higher (P<0.05) in the acute phase, as well as the inflammatory mediators (NOx, TNF-α, INF-γ, IL-4, IL-6) when compared to the control group. On the other hand, IL-10 levels were lower in the acute phase. Based on these results, we are able to conclude that the acute infection caused by E. canis in dogs leads to an increase on seric BChE activity and some inflammatory mediators. Therefore, this enzyme might be used as a marker of acute inflammatory response in dogs naturally infected by this bacterium.

Anaplasma phagocytophilum infection in a horse from Switzerland with severe neurological symptoms.

A 22-year old mare from Switzerland was admitted to an equine clinic in May 2011. She presented with fever, lethargy, icteric mucous membranes, reduced alertness, an unsteady gait and ataxia. An Anaplasma phagocytophilum infection was confirmed by blood smear and PCR. The mare was treated with oxytetracylin and recovered rapidly, but she still suffered from a slight atactic gait disturbance at 3 weeks post infection.

Acute respiratory distress syndrome in human granulocytic ehrlichiosis.

The ehrlichioses are emerging zoonotic infections. They have a very nonspecific clinical presentation. The diseases generally present as undifferentiated fever, but thrombocytopenia, leucopenia, and transaminitis are important laboratory features. Ehrlichiosis as a cause of acute respiratory distress syndrome (ARDS) has been infrequently described in the literature. Physicians should be aware of this life-threatening but treatable entity. We present a patient who developed ARDS shortly after being diagnosed with and initiating treatment for human granulocytic ehrlichiosis (HGE). A 54-year-old woman presented with fever, hypotension, and pancytopenia, with peripheral smear diagnostic of ehrlichiosis. She was started on doxycycline therapy following which she developed ARDS which resolved with methylprednisone. The fatality for HGE has been estimated as 7-10%. ARDS secondary to Ehrlichia has been shown to respond dramatically to steroids.

Lessons from Anaplasma phagocytophilum: chromatin remodeling by bacterial effectors.

Bacterial pathogens can alter global host gene expression via histone modifications and chromatin remodeling in order to subvert host responses, including those involved with innate immunity, allowing for bacterial survival. Shigella flexneri, Listeria monocytogenes, Chlamydia trachomatis, and Anaplasma phagocytophilum express effector proteins that modify host histones and chromatin structure. A. phagocytophilum modulates granulocyte respiratory burst in part by dampening transcription of several key phagocyte oxidase genes. The A. phagocytophilum protein AnkA localizes to the myeloid cell nucleus where it binds AT-rich regions in the CYBB promoter and decreases its transcription. AT-rich regions of DNA are characteristic of matrix attachment regions (MARs) which are critical for chromatin structure and transcription. MAR-binding proteins, such as SATB1, interact with histone modifying enzymes resulting in altered gene expression. With A. phagocytophilum infection, histone deacetylase 1 (HDAC1) expression is increased and histone H3 acetylation is decreased at the CYBB promoter, suggesting a role for AnkA in altering host epigenetics and modulating gene transcription, at this, and perhaps other loci. This review will focus on how bacterial pathogens alter host epigenetics, by specifically examining A. phagocytophilum AnkA cis-regulation of CYBB transcription and epigenetic changes associated with infection.

Epidemiology of ehrlichiosis and anaplasmosis among American Indians in the United States, 2000-2007.

Ehrlichiosis and anaplasmosis infections among American Indians (AIs) have never been specifically examined, despite high rates of other tick-borne rickettsial diseases among AIs. The epidemiology of ehrlichiosis and anaplasmosis among AIs was analyzed using the National Electronic Telecommunications System for Surveillance (NETSS), Case Report Forms (CRFs), and Indian Health Service (IHS) inpatient and outpatient visits. The 2000-2007 average annual ehrlichiosis and anaplasmosis incidence among AIs reported to NETSS was almost 4-fold lower (4.0/1,000,000) than that using IHS data (14.9). American Indian cases reported from CRFs had a higher proportion of hospitalization (44%) compared with IHS (10%). American Indian incidence of ehrlichiosis and anaplasmosis was higher and showed a different age and geographical distribution than other races. These results highlight the need to improve collaboration between the ehrlichiosis and anaplasmosis surveillance systems for AIs so as to develop interventions that target the unique epidemiology and mitigate the burden of disease among this high-risk population.

Ehrlichia chaffeensis presenting with bilateral anterior thigh pain (Louria's sign).

Bilateral anterior thigh pain may indicate bacteremia (Louria's Sign). We present a case of Ehrlichiosis due to Ehrlichia chaffeensis whose predominant presenting symptom was localized bilateral anterior thigh pain.

Experimental primary and secondary infections of domestic dogs with Ehrlichia ewingii.

In this study, the infection dynamics of Ehrlichia ewingii, causative agent of granulocytotropic ehrlichiosis in dogs and humans, was examined in experimentally infected dogs by using a combination of physical examination, hematologic and biochemical analyses, and molecular and serologic assays. For the experimental trials, blood from an E. ewingii-infected dog was inoculated intravenously into two naïve dogs and two dogs with prior experimental exposure to E. ewingii (both were negative for E. ewingii DNA by polymerase chain reaction (PCR) assay, but seropositive from initial infection 8 and 10 months prior to challenge). A negative control dog was inoculated with blood from a negative dog. The two primary infection dogs were positive for E. ewingii DNA on DPI 4, remained consistently positive until DPI 60, and were intermittently positive until the end of the study (DPI 144). The two primary infection dogs developed antibodies reactive to E. ewingii by DPI 28 and remained seropositive for the duration of the study. Primary infected dogs had intermittent fever, thrombocytopenia, and leukopenia and some dogs were hyperphosphatemic and/or had elevated ALP levels. The two challenge dogs were positive for E. ewingii DNA on DPI 4 and 18, which was similar to the primary infection dogs, but the duration of E. ewingii DNA detection was shorter. Also, the two challenged dogs did not develop pyrexia or show any hematologic or biochemical abnormalities. E. ewingii was successfully transmitted between dogs by Amblyomma americanum, but not Rhipicephalus sanguineus. This study provides data on the infection dynamics of E. ewingii in dogs during primary and challenge infections and suggests that prior exposure may lessen clinical disease during subsequent infections.

Variant -and individual dependent nature of persistent Anaplasma phagocytophilum infection.

BACKGROUND: Anaplasma phagocytophilum is the causative agent of tick-borne fever in ruminants and human granulocytotropic anaplasmosis (HGA). The bacterium is able to survive for several months in immune-competent sheep by modifying important cellular and humoral defence mechanisms. Little is known about how different strains of A. phagocytophilum propagate in their natural hosts during persistent infection. METHODS: Two groups of five lambs were infected with each of two 16S rRNA gene variants of A. phagocytophilum, i.e. 16S variant 1 which is identical to GenBank no M73220 and 16S variant 2 which is identical to GenBank no AF336220, respectively. The lambs were infected intravenously and followed by blood sampling for six months. A. phagocytophilum infection in the peripheral blood was detected by absolute quantitative real-time PCR. RESULTS: Both 16S rRNA gene variants of A. phagocytophilum established persistent infection for at least six months and showed cyclic bacteraemias, but variant 1 introduced more frequent periods of bacteraemia and higher number of organisms than 16S rRNA gene variant 2 in the peripheral blood. CONCLUSION: Organisms were available from blood more or less constantly during the persistent infection and there were individual differences in cyclic activity of A. phagocytophilum in the infected animals. Two 16S rRNA gene variants of A. phagocytophilum show differences in cyclic activity during persistent infection in lambs.

Current management of human granulocytic anaplasmosis, human monocytic ehrlichiosis and Ehrlichia ewingii ehrlichiosis.

Anaplasma phagocytophilum, Ehrlichia chaffeensis and Ehrlichia ewingii are emerging tick-borne pathogens and are the causative agents of human granulocytic anaplasmosis, human monocytic ehrlichiosis and E. ewingii ehrlichiosis, respectively. Collectively, these are referred to as human ehrlichioses. These obligate intracellular bacterial pathogens of the family Anaplasmataceae are transmitted by Ixodes spp. or Amblyomma americanum ticks and infect peripherally circulating leukocytes to cause infections that range in clinical spectra from asymptomatic seroconversion to mild, severe or, in rare instances, fatal disease. This review describes: the ecology of each pathogen; the epidemiology, clinical signs and symptoms of the human diseases that each causes; the choice methods for diagnosing and treating human ehrlichioses; recommendations for patient management; and is concluded with suggestions for potential future research.

Type IV secretion system of Anaplasma phagocytophilum and Ehrlichia chaffeensis.

The intracellular bacterial pathogens Ehrlichia chaffeensis and Anaplasma phagocytophilum have evolved to infect leukocytes and hijack biological compounds and processes of these host defensive cells. Bacterial type IV secretion (T4S) system transports macromolecules across the membrane in an ATP-dependent manner and is increasingly recognized as a virulence factor delivery mechanism that allows pathogens to modulate eukaryotic cell functions for their own benefit. Genes encoding T4S system homologous to those of a plant pathogen Agrobacterium tumefaciens have been identified in E. chaffeensis and A. phagocytophilum. Upon interaction with new host cells, E. chaffeensis and A. phagocytophilum genes encoding the T4S apparatus are upregulated. The delivered macromolecules are referred to as T4S substrates, or effectors, because they affect and alter basic host cellular processes, resulting in disease development. Recently, A. phagocytophilum 160-kDa AnkA protein was to be delivered by T4S system into the host cytoplasm. Thus, dynamic signal transduction events are likely induced by T4S substrates in the host cells for successful establishment of intracellular infection. Further studies on Ehrlichia and Anaplasma T4S effectors cognate host cell molecules will undoubtedly advance our understanding of the complex interplay between obligatory intracellular pathogens and their hosts. Such data can be applied toward treatment, diagnosis, and control of ehrlichiosis and anaplasmosis.

Clinical outcomes of solid organ transplant recipients with ehrlichiosis.

Because of our experience with severe Ehrlichia infections in lung transplant recipients, we reviewed all cases of ehrlichiosis in solid organ transplant recipients at Barnes-Jewish Hospital in St. Louis, Missouri. Between 1996 and 2007, 25 cases of ehrlichiosis were identified. We retrospectively collected demographic, clinical, laboratory, and outcomes data, and we compared the 5 cases in lung transplant recipients with 20 cases in other solid organ transplant recipients (heart, 2; kidney, 13; liver, 5). The presenting symptoms in the majority of both groups consisted of fever and headache. Clinical outcomes were worse in the lung transplant group and included a greater need for intensive care unit treatment (80% vs. 20%, P=0.02), longer length of hospital stay (21 vs. 5 days, P=0.02), and propensity to develop acute lung injury or acute respiratory distress syndrome (60% vs. 10%, P=0.04). No mortalities occurred in either group of patients. In an endemic area, ehrlichiosis is not unusual in solid organ transplant recipients, and lung transplant recipients tend to have a more severe illness.

Human granulocytic ehrlichiosis complicating early pregnancy.

BACKGROUND: The goal of this case is to review the zoonotic infection, human granulocytic ehrlichiosis, presenting with pyrexia. Case. A 22-year-old multigravid female presented to the emergency department with a painful skin rash, high fever, and severe myalgias. The patient underwent a diagnostic evaluation for zoonotic infections due to her geographical and seasonal risk factors. Treatment of human granulocytic ehrlichiosis was successful though the patient spontaneously aborted presumably due to the severity of the acute illness. CONCLUSION: Treatment of human granulocytic ehrlichiosis in pregnancy presents unique challenges. Management of pyrexia during pregnancy is limited to external cooling in the setting of thrombocytopenia and elevated aminotransferases. Extensive counseling regarding teratogenic potential of medications allows the patient to weigh the pros and cons of treatment.

Associations between proteinuria, systemic hypertension and glomerular filtration rate in dogs with renal and non-renal diseases.

Proteinuria and systemic hypertension are well recognised risk factors in chronic renal failure (CRF). They are consequences of renal disease but also lead to a further loss of functional kidney tissue. The objectives of this study were to investigate the associations between proteinuria, systemic hypertension and glomerular filtration rate (GFR) in dogs with naturally occurring renal and non-renal diseases, and to determine whether proteinuria and hypertension were associated with shorter survival times in dogs with CRF. Measurements of exogenous creatinine plasma clearance (ECPC), urine protein:creatinine ratio (UPC), and Doppler sonographic measurements of systolic blood pressure (SBP) were made in 60 dogs with various diseases. There was a weak but significant inverse correlation between UPC and ECPC, a significant inverse correlation between SBP and ECPC and a weak but significant positive correlation between UPC and SBP. Some of the dogs with CRF were proteinuric and almost all were hypertensive. Neoplasia was commonly associated with proteinuria in the dogs with a normal ECPC. CRF was the most common cause leading to hypertension. In the dogs with CRF, hypertension and marked proteinuria were associated with significantly shorter survival times.

Sequential analysis of Anaplasma phagocytophilum msp2 transcription in murine and equine models of human granulocytic anaplasmosis.

Anaplasma phagocytophilum causes human granulocytic anaplasmosis by inducing immunopathologic responses. Its immunodominant Msp2 protein is encoded by a family of >100 paralogs. Msp2 (msp2) expression modulates in the absence of immune pressure, and prolonged in vitro passage modulates in vivo virulence. Because programmed MSP2 expression occurs in Anaplasma marginale, we hypothesized a similar event in A. phagocytophilum in vivo, with specific Msp2 expression triggering immunopathologic injury or clinical manifestations of disease. We examined msp2 transcripts in 11 B6 mice and 6 horses inoculated with low- or high-passage A. phagocytophilum Webster strain. Blood was sequentially obtained through 3 weeks postinfection for msp2 reverse transcription-PCR. Horses were additionally assessed for clinical manifestations, seroconversion, complete blood count, blood chemistry, and cytokine gene transcription. In both species, there was no consistent emergence of msp2 transcripts, and all 22 msp2 variants were detected in both passage groups. Clinical severity was much higher for high-passage-infected than for low-passage-infected horses, preceded by higher levels of blood gamma interferon transcription on day 7. Antibody was first detected on day 7, and all horses seroconverted by day 22, with a trend toward lower antibody titers in low-passage-infected animals. Leukocyte and platelet counts were similar between experimental groups except on day 13, when low-passage-infected animals had more profound thrombocytopenia. These findings corroborate studies with mice, where msp2 diversity did not explain differences in hepatic histopathology, but differ from the paradigm of low-passage A. phagocytophilum causing more significant clinical illness. Alteration in transcription of msp2 has no bearing on clinical disease in horses, suggesting the existence of a separate proinflammatory component differentially expressed with changing in vitro passage.